An update on the biophysical character of the human eukaryotic elongation factor 1 beta: Perspectives from interaction with elongation factor 1 gamma.

ABSTRACT

The ß-subunit of the human eukaryotic elongation factor 1 complex (heEF1ß) plays a central role in the elongation step in eukaryotic protein biosynthesis, which essentially involves interaction with the α- and γ-subunits (eEF1γ). To biophysically characterize heEF1ß, we constructed 3 Escherichia coli expression vector systems for recombinant expression of the full length (FL-heEF1ß), N-terminus (NT-heEF1ß), and the C-terminus (CT-heEF1ß) regions of the protein. Our results suggest that heEF1ß is predominantly alpha-helical and possesses an accessible hydrophobic cavity in the CT-heEF1ß. Both FL-heEF1ß and NT-heEF1ß form dimers of size 62 and 30 kDa, respectively, but the CT-heEF1ß is monomeric. FL-heEF1ß interacts with the N-terminus glutathione transferase-like domain of heEF1γ (NT-heEF1γ) to form a 195-kDa complex or a 230-kDa complex in the presence of oxidized glutathione. On the other hand, NT-heEF1ß forms a 170-kDa complex with NT-heEF1γ and a high molecular weight aggregate of size greater than 670 kDa. Surface plasmon resonance analysis confirmed that (by fitting the Langmuir 11 model) FL-heEF1ß associated with monomeric or dimeric NT-heEF1γ at a rapid rate and slowly dissociated, suggesting strong functional affinity (KD  = 9.6 nM for monomeric or 11.3 nM for dimeric NT-heEF1γ). We postulate that the N-terminus region of heEF1ß may be responsible for its dimerization and the C-terminus region of heEF1ß modulates the formation of an ordered heEF1ß-γ oligomer, a structure that may be essential in the elongation step of eukaryotic protein biosynthesis.